Inflammatoric processes, such as sepsis, are a major cause of morbidity and mortality in humans. It is estimated that, yearly, 400 000 to 500 000 episodes of sepsis results in 100 000 to 175 000 human deaths in the U.S. alone. In Germany, sepsis rates of up to 19% of patients stationed at Intensive Care Units have been noted. Sepsis has also become the leading cause of death in intensive care units among patients with non-traumatic illnesses. Despite the major advances of the past decades in the treatment of serious infections, the incidence and mortality due to sepsis continues to rise.
There are three major types of sepsis characterized by the type of infecting organism. Gram-negative sepsis is the most common. The majority of these infections are caused by Esherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. Gram-positive pathogens, such as the staphylococci and the streptococci, are the second major cause of sepsis. The third major group includes the fungi. Fungal infections constitute a relatively small percentage of the sepsis cases, but they result in a high mortality rate.
A well-established mechanism in sepsis is related to the toxic components of gram-negative bacteria, i.e. the lipopolysaccharide cell wall structure (LPS, endotoxin), which is composed of a fatty acid group, a phosphate group, and a carbohydrate chain.
Several of the host responses to endotoxins have been identified, such as release of cytokines, which are produced locally. In case of an extensive stimulation, however, there is a spill over to the peripheral blood and potential harmful effects are obtained, such as induced organ dysfunction.
The key mediators of septic shock are Tumor Necrosis Factor (TNF-α), Interleukine 1 (Il-1) and Interleukine 17 (Il-17), which are released by monocytes and macrophages. They act synergistically causing a cascade of physiological changes leading to circulation collapse and multi organ failure. Indeed, high concentrations of TNF-α can mimic the symptoms and outcome of sepsis.
Normally, endotoxins are kept within the lumen of the intestine. For example, during cardiopulmonary bypass the presence of splanchic ischemia or dysoxia causes disruption of the mucosal barrier and translocation (i.e. the transport of endotoxins from the intestine to the circulation system) of endotoxins from the gut lumen to the portal circulation.
Antibiotics of varying types are widely used to prevent and treat infections. However, for many commonly used antibiotics an antibiotic resistance is developed among various species of bacteria. This is particularly true for the microbial flora resident in hospitals, where organisms are under a constant selective pressure to develop resistance. Furthermore, in the hospital the high density of potentially infected patients and the extent of staff-to-staff and staff-to-patient contact facilitate the spreading of antibiotic resistant organisms. The antibiotics used are the most economical, the safest and the most easy to administer and may not have a broad enough spectrum to suppress certain infections. Antibiotics can be toxic to varying degrees by causing allergy, interactions with other drugs, and causing direct damage to major organs (e.g. liver, kidney). Many antibiotics also change the normal intestinal flora, which can cause diarrhea and nutritional malabsorption.
Certain antibiotics are known to neutralize the action of endotoxins, such as polymyxin B. This antibiotic binds to the lipid A part of endotoxin and neutralizes its activity. Polymyxin is not used routinely due to its toxicity. It is only given to patients under constant supervision and monitoring of the renal function.
Furthermore, in order to overcome some of the limitations inherent to active immunization against bacterial components, various techniques have been used to produce endotoxin-binding antibodies. A large number of antibodies have been prepared by immunization of humans with bacteria. In order to develop more consistent preparations of therapeutic antibodies, numerous LPS-reactive monoclonal antibodies have been developed. Unfortunately, the clinical studies have not resulted in benefits. However, it should be noted that these trials were performed in humans after onset of symptoms of sepsis. It is widely believed that an anti-endotoxin antibody treatment, administered after sepsis, may yield little benefit because these antibodies cannot reverse the inflammatory cascade initiated by the endotoxin.
In JP 06022633, an adsorbent for anti-lipid antibodies is shown, which comprises a compound with an anionic functional group immobilized onto a water-insoluble porous material. The porous material can be agarose, cellulose, dextran, polyacrylamide, glass, silica gel, or a hard polymer made of a styrene-divinylbenzene copolymer, and the porous material is packed as a bed of separate particles in a separation device.
In attempts to remove components from blood, different adsorbent materials have been prepared. An endotoxin removal adsorbent comprising a ligand immobilized on a solid phase support medium is shown in WO 01/23413. A preferred support medium is in the form of beads. When packed in a separation device, the solid phase support medium is porous enough to allow passage of blood cells between the beads.
In WO 00/62836, the adsorbent material has a size and a structure adapted to remove β-2 microglobulin from blood. The adsorbent material of this document can be a macroporous synthetic polymer with a surface of beads and of pores modified as to prevent adsorption of proteins and platelets. However, individual spherical beads of the polymer were mechanically destroyed at a loading of about 500 g, which is obtained in for example a column packed with the beads. Such a loading results in a considerable pressure drop over of the column.
In order to reduce the pressure drop, an absorbent has been prepared in EP 464872, which comprises water-insoluble porous hard gel particles having an exclusion limit of 106-109 Dalton. The gel bed is used for selective removal of lipoproteins from blood or plasma in extra-corporeal circulation treatment.
Likewise, in WO 01/23413 the porous support material for endotoxin removal is beads, which can be filled into a container, the beads having a size sufficient to provide the requisite space between the beads when packed into a column or filter bed. The porous support material can also be microfiltration hollow-fibers or flat sheet membranes in order to minimize pressure drops.
In EP 424698 an adsorbent for eliminating biomacro-molecules is shown, which consists of a carrier of porous spherical particles having a particle size of 50-150 microns and an exclusion limit of at least 105 Dalton. Polymyxin B is coupled to the particles, which are subsequently filled in a cartridge to be used in a system for extracorporeal endotoxin removal from whole blood.
In these traditional systems for extracorporeal removal of toxic components from blood, a container or cartridge is first filled with a liquid and the adsorbing porous beads are introduced afterwards. In U.S. Pat. No. 6,408,894 a method is shown, which provides a more uniform distribution and denser packing of the beads. The method involves forcedly supplying a mixture of liquid and beads into a container in such a manner that the liquid is squeezed out of the mixture and out of the container.
Thus, an elimination of blood cells facilitates the removal of compounds present in plasma as described above, e.g. in WO 00/62836 or WO 01/23413. However, such a technique involves two separation steps which both could contribute to an enhanced risk of adverse cellular activation due to bioincompatability.